Stage One,Brainstorming for Project
During our brainstorming sessions for the project, the team proposed a myriad of research topics. However, one particular area that piqued everyone's interest was the environmental impact of insecticides. A common insect repellent that most of us frequently use in our daily lives is mosquito repellent. After thorough literature research, we discovered that the primary ingredient in many of these repellents is pyrethroids, a synthetic chemical similar to the natural pyrethrins produced by chrysanthemum flowers. Recognizing the ubiquity of this compound and its potential ecological implications, we decided to delve deeper into understanding its effects and role in the environment.
Interview with Professor Xue from Zhongyang Minzu Universit
Objective: To Understand the Environmental Impact of Pyrethroids
>In our quest to understand the environmental ramifications of pyrethroids, a common ingredient in many insect repellents, our team went beyond mere literature reviews. We sought the expertise of Professor Xue from the National University, a renowned entomologist. During our insightful interview, Professor Xue shared findings from his recent research where he attempted to identify microbial communities in the gut of wild insects that might resist insecticides. Despite his rigorous investigations, there was no substantial evidence supporting the existence of such resistant microbes. He emphasized that insecticides like pyrethroids pose significant threats not just to pests, but also to beneficial and neutral insects in the wild, including pollinators and decomposers. This interaction with Professor Xue reinforced the value of our project, underscoring the urgent need for sustainable solutions. His insights significantly shaped our approach, steering us towards a more comprehensive understanding of the broader ecosystem impacts of insecticides.
Interview with Professor Yao from the China Central Bee Museum
Objective: Understand which impacted insect is most worthy of attention
Outcome: Identified bees as our primary insect of concern
We had the privilege of interviewing Professor Yao, the Director of the China Central Bee Museum and a distinguished member of the Bee Research Institute under the Chinese Academy of Agricultural Sciences. During our insightful discussion, Professor Yao delved into the intricate relationship between bees, pesticides, and agricultural ecology. He highlighted that bees not only represent a species whose impact, when affected, results in significant economic ramifications, but they also play a pivotal role in contributing to biodiversity. Emphasizing the immense value of bees in the ecosystem, he strongly encouraged our team to focus our attention and efforts on them. This engagement with Professor Yao underscored the importance and relevance of our project, offering a more directed approach. His expertise greatly influenced our perspective, urging us to prioritize bees as our primary insect of concern, thereby shaping the trajectory of our research endeavors.
Interview with Water Environment Expert, Professor Hao
Objective: Understand the primary ways in which pyrethroids affect bees
Outcome: Established addressing water pollution as the project's objective
During our interview with Professor Hao, an expert in water environments, we gained a deeper understanding of the primary pathways through which pesticide pollution spreads. Of particular note was the way pesticides, via water pollution, impact wild insects. Common household items such as insect sprays, mosquito coils, and airborne residues might find their way into water systems through rain and eventually end up in wastewater treatment plants. Specifically, pyrethroids residues, a main component in many insecticides, can enter these water systems simply through routine kitchen cleaning.
Given this information, we decided to position our project towards purifying wastewater of pyrethroids, aiming to mitigate environmental harm. In tandem, we embarked on creating informational brochures to raise public awareness about the responsible use of pesticides and the importance of protecting our delicate ecosystems. This engagement with Professor Hao not only validated the urgency and value of our project but also offered a clear direction, emphasizing the intersection of community awareness and innovative solutions for environmental restoration.
Creating a Brochure: Highlighting the Potential Hazards of Pyrethroids and Guidelines for Safe Use
Our feedback: Using synthetic biology to produce astragalin
We've developed an informative flyer designed to enlighten its readers on several key topics central to our project. Firstly, it introduces the basics of enzymes, detailing their nature and importance in biological processes. It also educates the general public of synthetic biology, elucidating its principles and its transformative potential in modern science and technology. Lastly, the flyer highlights the harmful effects of pyrethroids, aiming to raise awareness about their potential hazards and the significance of their responsible use. This flyer serves as a inspiring guide for people to restrict the usage of pyrethroid when necessary.
Stage TWO，Engineering for the Project
Literature Research: Objective: Determine the degradation methods for pyrethroids Outcome: Identified 5 target enzymes
Through our comprehensive literature research, we identified four enzymes with promising expression results. Our immediate plan encompasses studying the production expression of these four enzymes to better understand their potential in addressing the issue of pyrethroids degradation. In the course of our research, we also took note of a prior project by the iGEM 2022 Warwick team who similarly focused on the pyrethroids issue. Their scope was limited to the study of a single enzyme and its efficiency against a specific pyrethroid. Recognizing the value of building upon existing research, we have chosen to compare the efficacy of our identified enzymes with the enzyme studied by the Warwick team. This comparative approach not only validates the importance of our project but also helps refine our research direction, ensuring a holistic understanding of pyrethroids degradation. Moreover, by leveraging the insights from a previous iGEM team, our research can contribute to a continuous and collaborative knowledge-building process in this domain.
An example of the coding sequencing EstA embedded into an expression vector pET-28a.
Interview with Biology Experiment Expert, Dr. He
Objective: Enhance experimental proficiency.
Outcome: Recommended utilizing HPLC for quantitative analysis
In our pursuit of advancing our experimental methodologies, we sought the expertise of Dr. He, a renowned biology experiment expert from the GenTex Biotechnology Institute. Our primary goal during this interview was to gather insights that would elevate our experimental proficiency. Dr. He's valuable recommendation was to adopt High-Performance Liquid Chromatography (HPLC) for quantitative analysis. The adoption of HPLC, as suggested by Dr. He, offers precise and reliable results, which are essential for the integrity of our project. This recommendation was instrumental in refining our research approach, ensuring that our data collection methods are both accurate and efficient. The guidance from an expert like Dr. He underscores the value of our project, and emphasizes the importance of integrating advanced techniques to achieve our objectives. By incorporating his advice, our project stands to benefit from robust experimental data, paving the way for more informed conclusions and potential applications.
Interview with Bio-experiment Expert, Dr. Liu Huan from BGI (Beijing Genomics Institute)
Objective: Finalize details of the experimental detection design
Outcome: Decided to use the paper strip method to test enzyme degradation efficiency
After considerations of the use of HPLC, it became clear that we lacked the capability to utilize this specific detection method. Recognizing the necessity for an alternative, we sought expertise from Dr. Liu at BGI (Beijing Genomics Institute). Through our interview, Dr. Liu brought to our attention a paper strip method capable of detecting pyrethroids. He suggested that we could employ this paper strip as a qualitative measure for our project. This pivotal interaction underscored the adaptive nature of scientific research, where challenges often lead to innovative problem-solving. The recommendation from Teacher Liu not only provided a feasible solution to our initial challenge but also emphasized the resourcefulness required in the field of experimental research. By adopting this paper strip method, our project remains rooted in effective and applicable methodologies, ensuring that our research goals are met in a pragmatic and efficient manner. The invaluable guidance from experts like Teacher Liu reinforces the worth of our project, illustrating the importance of flexibility and collaboration in the scientific endeavor.
Dr. Liu Huan from BGI.
Interview with Professor Geng, Bio-experiment Expert from Zhongyang Minzu University
Objective: Confirm successful protein expression
Outcome: Professor Geng assisted with a Western Blot (WB) test, confirming successful results.
In our initial attempt to verify protein expression, we utilized SDS-PAGE. However, while the results did show protein bands, there were numerous distinct bands present, leading to uncertainty about the specific protein expression. Given this ambiguity, we sought the expertise of Professor Geng from Zhongyang Minzu University. With her assistance, a more sensitive Western Blot (WB) analysis was conducted. The results from the WB clearly confirmed the successful expression of our target protein.
This supporting evidence underscored the significance of leveraging advanced techniques and expert insights when faced with experimental uncertainties. Professor Geng's intervention was pivotal, helping us navigate challenges and ensuring that our research remained on the right track.
Prof. Geng showed us lab techniques for better experiment results.
SDS-Page Results of EstA
Visit to the Second Largest Water Treatment Plant in Tongzhou District of Beijing City
Objective: Research application scenarios and establish experimental conditions.
Outcome: Determined the optimal pH, temperature, and time range for enzyme production and activity.
During our comprehensive site visit to the Tongzhou Wastewater Treatment Plant in Beijing, we observed multiple stages of water purification. Among these stages, the aerobic fermentation phase held particular relevance for our project. In this phase, microbes in the wastewater ferment, aiding in the removal of organic pollutants. We noted that during the winter, the operational temperature range lies between 16-25°C, while in the summer, temperatures often exceed 35°C. The pH level remains approximately 7.1, and the wastewater stays in the purification system for 12-16 hours.
Drawing insights from our visit, we decided to conduct experiments at the specified temperature ranges and pH levels. We also planned to assess which enzyme exhibits optimal degradation efficiency within the 12-16 hour timeframe. This targeted approach, directly informed by our on-site observations, ensures that our project is both applicable and tailored to the specific conditions of the wastewater treatment plant, enhancing its potential value and impact.
Literature Review: Refinement of the 2022 Warwick Submission Components
Objective: Identify limitations and potential improvements in the genetic components submitted by 2022 Warwick team.
Outcome: Improved genetic circuitry leading to correct expression.
Initially, we utilized sequences available on the iGEM parts repository for our experiments. However, disappointingly, this approach did not result in any protein expression. Recognizing this challenge as an opportunity for further refinement, we revisited the literature meticulously. Instead of solely relying on the parts repository, we delved deep into the original research papers and reconstructed new sequences based on the foundational literature. After redesigning the sequences and expressing them anew, we were rewarded with successful protein expression results. This exercise underscored the importance of a diligent literature review and its invaluable role in troubleshooting and enhancing experimental outcomes. Moreover, this experience reinforced our belief in the iterative nature of scientific research, demonstrating that setbacks can often be stepping stones to more robust and validated results. By refining and validating our methods based on original sources, our project not only attained its immediate objectives but also contributed a more dependable sequence to the broader scientific community.
Sequence of the newly designed Pyre1.
Stage THREE,Applications for the Project
Interview with Dr. Perriman from the University of Bristol
Objective: Enhancing Application Efficiency
During our project's development, understanding the dynamics of enzyme activity in real-world settings became crucial. We sought expertise from Dr. Perriman, a renowned figure from the University of Bristol, known for his extensive work in biotechnology. He shed light on various strategies to optimize enzyme efficiency. These included environmental considerations such as pH, temperature, and substrate concentration, as well as protein engineering techniques to modify the enzyme's active site or enhance its stability. Additionally, he emphasized the importance of immobilizing enzymes on solid supports, which not only extends their functional lifespan but also facilitates their recovery and reuse – a pivotal aspect for industrial applications. This interaction with Dr. Perriman became a cornerstone for our project. By integrating his insights, we could fine-tune our approach, ensuring that our enzyme not only functioned optimally in lab conditions but also thrived in real-world environments, increasing the overall value and impact of our work.
Consultation with Dr. Ma, Technical Director of an Environmental Protection Company, and Expert Reviewer of Pesticide Production Licenses, Dr. Wei
Outcome: The project contributes significantly to biodiversity.
We had the opportunity to present our project to two experts: Dr. Ma, the Technical Director of an Environmental Protection Company, and Dr. Wei, an expert in pesticide production license reviews. Both recognized the significance of our project's contribution to biodiversity. Biodiversity is crucial for maintaining ecosystem resilience and providing essential services like food, clean water, and raw materials. Our project aims to reduce the negative effects of pesticide residues on non-target organisms, promoting a balanced ecosystem. The feedback from Dr. Ma and Dr. Wei confirmed the value of our work and its potential positive impact on the environment.
Interview with Professor Zhou from the College of Agriculture's Entomology Department
Outcome: Suggested considering the modification of bee gut microbiota based on our project, to further protect insects.
We had the opportunity to interview Professor Zhou Xin from the College of Agriculture's Entomology Department at China Agricultural University. Notably, he also served as the inaugural Executive Director of China's National Gene Bank. Professor Zhou expressed high regard for the students' chosen topics and their subsequent actions. Furthermore, he provided valuable insights, particularly concerning the bee gut microbiota, offering new directions and perspectives for our project. This interaction underscored the significance and relevance of our project in the broader academic and environmental context.
Modifying the bee gut microbiota can be approached in several ways. One common method is through the introduction of probiotics, specifically tailored bacterial strains that can benefit the bee's health and potentially degrade harmful substances like pesticides. Another approach might involve using prebiotics, substances that promote the growth of beneficial microorganisms already present in the bee's gut. On a more advanced note, synthetic biology techniques could be employed to engineer specific bacterial strains that offer enhanced benefits or target particular challenges faced by bees.
However, while these interventions promise benefits, they also bring up biosafety concerns. Introducing modified or foreign bacteria into the bee gut could inadvertently harm the delicate balance of the existing microbial community, potentially leading to unintended health consequences for the bees. Furthermore, there's always a risk that these introduced organisms might spread beyond the target population, affecting other insects or even broader ecosystems. Thus, before any large-scale implementation, thorough research and risk assessment are essential to ensure the safety and efficacy of these interventions.
Visit to Institute of Plant Research, China, and Director Zhao of Beijing Botanical Garden
Result: The project's treated water (final water) is now frequently used for irrigation of trees and flowers in specific areas, which is highly significant for the protection of biodiversity."
